FIGURE 1.

Design and strategy of generating a conditional pi4ka allele. A, homologous recombination in ES cells was used to generate ES cells heterozygous for the Pi4ka primary targeted allele. The structure of the Pi4ka allele in the region of exon 48 is presented, with exons indicated as blue boxes. The structure of the targeting construct is shown with the short and long homology arms (SA and LA, respectively), the exon 48-containing target region, the neomycin selection cassette, and loxP and FRT elements (yellow and green arrowheads, respectively). The positions of the NsiI restriction sites (N), probes (gray boxes), and band sizes indicative of the wild-type and targeted Pi4ka allele used in the Southern blot screening strategy are indicated. The resultant primary targeted allele and the structure of the conditional and null alleles are shown following, respectively, Flp and Cre recombinase-mediated deletion. B, representative Southern blots using the 5′ and 3′ probes indicated in A. Genomic DNA from targeted ES cell clones (lanes 1–5), parental ES cell control DNA (C), and molecular size markers (M). Expected band sizes of the wild-type and targeted allele are indicated.